Biocompatible sensor electrode assembly and method for the production thereof

ABSTRACT

The invention relates to a biocompatible sensor electrode arrangement and to a process for its manufacture, at least one carrier substrate area ( 22 ), at least one intermediate substrate area ( 26 ) on the surface area ( 22   a ) of the carrier substrate area ( 22 ) and a biomaterial area ( 24 ) on the top side surface area ( 26   a ) of the intermediate substrate area ( 26 ) being provided. The biomaterial area ( 24 ) consists of at least one biologically compatible material component. The carrier substrate area ( 22 ) with the intermediate substrate area ( 26 ) is formed in the form or the manner of a wafer element or a printed circuit, as photolithographically processed structure, as structure bonded on or laminated on and/or as structure processed by printing, in particular on the carrier substrate ( 22 ) in each case.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a National Stage application of InternationalApplication No. PCT/EP2004/004993, filed on May 10, 2004, which claimspriority of German application number 103 20 898.4, filed on May 9,2003.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The invention relates to a biocompatible or biologically compatiblesensor electrode arrangement and a process for its manufacture.

2. Description of the Prior Art

In many areas of chemical and biochemical analysis technology,biocompatible or biologically compatible material arrangements, inparticular biocompatible or biologically compatible sensor arrangementsor sensor electrode arrangements are used. As a result of these materialarrangements, sensor arrangements or sensor electrode arrangements,certain measuring processes are used in practical application withrespect to a chemical, biological or biochemically relevant analyte.

In the case of analytical processes with high rates of throughput, i.e.in the case of so-called high throughput screening processes, differentcharacteristics are desirable for biocompatible material arrangements,sensor arrangements or sensor electrode arrangements, in particular withregard to their electrical sensitivity, their mechanically stabilityand/or their high and cost-effective availability. In the case ofconventional material arrangements, sensor arrangements or sensorelectrode arrangements, carrier substrates are used which, althoughhaving a mechanically relatively stable construction, they may entail anotherwise comparatively difficult handleability and also not benecessarily producible in a cost-effect manner.

SUMMARY OF THE INVENTION

The invention is based on the task of indicating a biocompatible orbiologically compatible sensor electrode arrangement and a process forits manufacture in the case of which cost-effective and both reliablyhandleable biocompatible or biologically compatible materialarrangements are used in a particularly simple manner.

This task is achieved in the case of a biologically compatible orbiocompatible sensor electrode arrangement according to the presentinvention. Moreover, the task is achieved according to the invention inthe case of a process for the manufacture of a biologically compatibleor biocompatible sensor electrode arrangement. The biologicallycompatible or biocompatible sensor electrode arrangement according tothe invention exhibits at least one carrier substrate area which isformed with a top side with a surface area or a top side surface area.Moreover, an intermediate substrate area or a connecting substrate areais provided which is formed on the surface area of the carrier substratearea or a part thereof, in particular in a structured manner, and whichis formed with a top side facing away from the carrier substrate areawith a surface area or with a top side surface area. Finally, abiomaterial area is provided which is formed on the top side surfacearea of the intermediate substrate area or the connecting area or a partthereof, in particular in a structured manner, with at least onebiocompatible or biologically compatible material component. The carriersubstrate area with the intermediate substrate area or the connectingsubstrate area thereon or the intermediate substrate area or theconnecting substrate area as such and/or a part thereof in each case areformed, for example, in the form or in the manner of a wafer element ora printed circuit. Alternatively or additionally, the intermediatesubstrate area or the connecting substrate area are formed as a or witha photolithographically processed structure or as a or with aphotographically processed element. As a further alternative oradditionally, the intermediate substrate area or the connectingsubstrate area are provided as a or with a structure processed by beingbonded on or laminated on or as an or with an element processed by beingbonded on or laminated on. As a further alternative or additionally, theintermediate substrate area or the connecting substrate area are formedas a or with a structure processed by printing or as an or with anelement processed by printing. Moreover, alternatively or additionally,the intermediate substrate area or the connecting substrate area areformed as a or with a structure processed micromechanically and/or bylaser ablation or as an or with an element processed micromechanicallyand/or by laser ablation.

This takes place in particularly directly on the carrier substrate areain each case.

It is consequently a core idea of the present invention to form thebiocompatible sensor electrode arrangement according to the inventionbased on a carrier substrate area, between the carrier substrate areaand the biomaterial area, an intermediate substrate area or connectingsubstrate area being provided by way of at least one biocompatiblematerial component.

A further core idea of the present invention consists of the fact thatthe carrier substrate area with the intermediate substrate area or theconnecting substrate area or that the intermediate substrate area orconnecting substrate area as such or parts thereof are provided as awafer element, as a printed circuit, as a photolithographicallyprocessed structure, as a structure bonded or laminated on, as structureprocessed micromechanically and/or by laser ablation and/or as astructure processed by printing, in particular on the substrate carrierarea in each case.

As a result of this structure provided according to the invention,advantages arise compared with the state of the art to the effect thatlow material and manufacture costs, for example, arise in the case ofthe biocompatible sensor electrode arrangement according to theinvention because common mass manufacture techniques connected with thesuggested modes of formation and structures can be used during themanufacture.

Moreover, a corresponding mechanical strength of the biocompatiblesensor electrode arrangement according to the invention is obtainedautomatically with a high reliability in use and a high applicationflexibility, the aspect of miniaturisation of the structures of thebiocompatible sensor electrode arrangements of the invention beingthereby taken into consideration, on the basis of the correspondingprocess technology, sufficiently and in a reliable manner.

Although it is possible to produce the intermediate substrate area orthe connecting substrate area by epitaxial growing, by vapour depositionand/or sputtering in a particularly well controlled, well defined mannerand/or with planar surfaces, it is possible under certain circumstancesfor straight structures and processes for their manufacture andcorresponding techniques to offer themselves for reasons of costs and/orsimplification, in the case of which such a control of the areaproperties and surface properties of the intermediate substrate area orthe connecting substrate area is not possible, such as e.g. in the caseof application and/or structurisation by photolithography, bonding,lamination, printing, ablation, laser ablation or such like if a gooddefinition, controllability and/or planarity of the intermediatesubstrate area or the connecting substrate area or their surface are notimportant.

In the case of a preferred embodiment of the biocompatible sensorelectrode arrangement according to the invention it is provided that thecarrier substrate area exhibits a chemically inert, biologically inertand/or essentially electrically insulated material or is formed of sucha material. As a result of this measure, minimal interactions with thechemical or biological surroundings arise and the use as a carrier foran electrode offers itself in particular.

With a view to the multiple application possibilities, it may beadditionally or alternatively provided for the carrier substrate area toexhibit a mechanically flexible material or to be formed of such amaterial, in particular in the manner or in the form of a film. Thisthen allows the manufacture of, e.g., single use sensors for singleapplication, e.g. for diagnostic or clinical purposes or for use in thefield, in situ or at the point of care.

With the regard to the structurisation of the biocompatible materialarrangement as a sensor electrode arrangement, it is providedadditionally or alternatively in the case of another embodiment of thesensor electrode arrangement according to the invention that a layer ofan electrically conductive metal oxide, e.g. ITO or indium tin oxide isformed for the intermediate substrate area or for the connectingsubstrate area.

On the other hand, it may be possible, with respect to thestructuralisation of the biocompatible material arrangement as a sensorelectrode arrangement in the case of another embodiment of the sensorelectrode arrangement according to the invention to be additionally oralternatively provided that a metallic layer structure is formed on thetop side surface area of the carrier substrate area for the intermediatesubstrate area or for the connecting substrate area.

In this connection, it may be advantageous if the layer structure forthe intermediate substrate area or for the connecting substrate area isformed with or from at least one primary metal area arranged at bottommost, a subsequent auxiliary layer and an actual electrode layerarranged top most.

The auxiliary layer can serve, in particular, as an alloy and/ordiffusion barrier between the primary metal area and the actualelectrode layer such that an alloy formation as a result ofinterdiffusion of the materials of the primary metal area and the actualelectrode layer into or with each other is avoided. This maintains, forexample, the specificity of the actual electrode layer, in particular inthe case where a surface functionalisation or surface improvement of theactual electrode layer is involved.

The primary metal area can be formed with or of copper, for example.

The structurisation of the primary metal area can be provided indifferent forms, e.g. in the form of a photolithographically structuredor applied primary metal area. In addition or alternatively, primarymetal areas applied by bonding, lamination, ablation and/or processed byprinting are conceivable.

According to another embodiment of the sensor electrode arrangementaccording to the invention, the auxiliary layer is formed with or fromnickel.

In the case of a further alternative embodiment of the sensor electrodearrangement according to the invention it is provided that the actualelectrode layer arranged top most is formed with or from a noble metal.Gold is preferably used in this respect.

The auxiliary layer and/or the actual electrode layer arranged top mostcan be formed by electrodeposition.

Alternatively or additionally, it is possible to consider amicromechanical formation and/or a laser ablation.

With respect to the further biological, chemical or biochemicalcompatibility of the sensor electrode arrangement as a whole, it can beprovided that the carrier substrate area is formed entirely or partlyfrom a chemically inert, biologically inert material and/or materialwith an at most lost adsorptivity vis-à-vis proteins, biological and/orchemical active substances.

With regard to the material selection, entirely different materials canbe used in the carrier substrate area:

It is conceivable to use glass, quartz and/or mica as materials for orin the carrier substrate area because, then, a particularlywell-defined, well-controlled and/or planar carrier substrate areaand/or surface area is then present.

However, it is advantageous if the carrier substrate area is formedentirely or partly of PMMA, PEEK, PTFE, POM, FR4 polyimide such as e.g.PI or Kapton, PEN, PET and/or a material which is transparent—inparticular in the UV range—if good definition, control and/or planarityof the carrier substrate area and/or the surface area is not important.As a result of this material selection, the processing effort and thecosts can be reduced.

The meaning in this respect is as follows: PMMA polymethylmethacrylatePEEK polyetheretherketone PEN polyethylene naphthalat FR4 glassfibre-reinforced epoxy resin PI polyimide PET polyethylene terephthalatePOM polyoxymethylene and PTFE polytetrafluoroethylene or Teflon

PEEK, POM and PTFE are not transparent as a rule. PTFE can usually notbe bonded. However, if bonding techniques other than UV bonding areused, these materials can also be used in a meaningful manner.

With respect to as high a surface yield as possible and/or possibleautomation, it is, moreover, advantageous if a multiplicity, inparticular of identical intermediate substrate areas or connectingsubstrate areas and/or corresponding biomaterial areas is formed. Thisplurality can be formed in a connected or in a separated form. Inparticular, an electrical insulation from each other offers itself inorder to obtain sensor elements of the sensor electrode arrangementwhich are separated and insulated from each other, for example. Themajority of intermediate substrate areas or connecting substrate areasand/or biomaterial areas is formed also in a form laterally arrangedside by side on the carrier substrate area, for example.

An arrangement of the plurality of intermediate substrate areas orconnecting substrate areas and/or corresponding biomaterial areas in asequence or matrix form is particularly advantageous.

In a particularly advantageous manner, the use of the sensor electrodearrangement according to the present invention offers itself as sensorelectrode arrangement for amperometric and/or potentiometricpharmacological effective site and/or active principle testing.

The intermediate substrate area and/or the biomaterial area or theircombination can be provided in an advantageous manner as membranebiosensor electrode area or as secondary carrier of the sensor electrodearrangement.

The intermediate substrate area or the connecting substrate area and thebiomaterial area can be formed respectively as a membrane biosensorelectrode area or as secondary carrier with an electrically conductiveand solid body-type electrode area.

With respect to the use in the area of amperometric and/orpotentiometric pharmacological active site and/or active principletesting, in particular, further aspects may be essential as will beexplained below.

In the case of the biocompatible or biologically compatible sensorarrangement or sensor electrode arrangement according to the inventionfor amperometric and/or potentiometric, pharmacological active siteand/or active principle testing, a membrane biosensor electrode area isprovided which, according to the meaning of the invention, is alsoreferred to synonymously as secondary carrier. This membrane biosensorelectrode area or secondary carrier exhibits an electrically conductiveand solid body-type electrode area.

This is formed by the intermediate substrate area or the connectingsubstrate area, for example, and in particular by the actual electrodelayer.

Moreover, a plurality of primary carriers is provided which are arrangedin the immediate spatial vicinity of the membrane biosensor electrodearea or the secondary carrier and which exhibit units which areactivable to an electrical action and biological, in particular membraneproteins. In addition, an aqueous measuring medium is provided in whichthe primary carriers and at least part of the membrane biosensorelectrode area or the secondary carrier are arranged.

According to the invention, the electrode area is formed in a mannerthat is electrically insulated vis-à-vis the measuring medium, theprimary carriers and vis-à-vis the biological units.

According to the invention, a eukaryotic cell, a prokaryotic cell, abacterium, a virus or components, in particular membrane fragments orassociations thereof in the native form or in the modified form, inparticular in the purified, microbiological form and/or form modified bymolecular biology are provided as primary carrier in each case.Alternatively or additionally, a vesicle, a liposome or a micellarstructure are provided as primary carrier.

An essential component of the sensor arrangement or sensor electrodearrangement according to the invention also consists of a membranebiosensor electrode area or a secondary carrier which, in the following,can also be called sensor electrode device. This sensor electrode devicefor amperometric and/or potentiometric, pharmacological active siteand/or active principle testing itself thus exhibits at least oneelectrically conductive electrode area. The sensor electrode device isformed such as to be arranged in an aqueous measuring medium duringoperation. Moreover, the sensor electrode device is formed such that aplurality or a multiplicity of primary carriers with biological unitsactivable into electrical action, in particular membrane proteins orsuch like are arranged in immediate spatial vicinity, in particular ofthe electrode area. According to the invention, at least the electrodearea is in this respect formed like a solid body. Moreover, according tothe invention, the electrode area is formed such that it is electricallyinsulated vis-à-vis the measuring medium to be provided and vis-à-visthe primary carriers.

It is thus a further idea of the present invention to form at least theelectrode area of the sensor arrangement according to the invention and,in particular, the sensor electrode device as a solid body or in a solidbody supported manner. As a result, the sensor electrode device and, inparticular, the provided electrode area thereof are provided with aparticularly high mechanical stability as a result of which aparticularly robust operation not susceptible to interference ispossible within the framework of the active site and/or active principletesting.

Only by solid body support is an activation e.g. of membrane proteinsmade possible by a concentration jump. This can take place, inparticular, within the framework of a rapid and/or continuous solutionexchange as a result of which—particularly in the case of amperometricmeasurements—a high signal level and consequently a high sensitivity canbe achieved. As a result of the robustness due to the solid bodysupport, easier handling and convenient incorporation are possible.

A further aspect of the present invention consists of forming theelectrode area in such a way that it is formed in an electricallyinsulated manner vis-à-vis the measuring medium and vis-à-vis theprimary carriers during operation. As a result of this measure it ispossible to use the sensor electrode device as a capacitively coupledelectrode, for example. This has considerable advantages particularlywith regard to the signal-to-noise ratio, i.e. with regard to theaccuracy of detection. Moreover, in the case of capacitive coupling, theelectrode area of the sensor electrode device does not participate inany chemical conversion such as would be the case with a typicalelectrochemical half-cell.

The selection of the primary carriers carrying the biological unitsneeds to be regarded as a further core aspect of the sensor arrangementaccording to the invention. The primary carriers may consist ofeukaryotic cells, prokaryotic cells, bacteria, viruses or components, inparticular membrane fragments or associations thereof respectively,namely in the native form or in a modified form, in particular in apurified form or a form modified by molecular biology and/ormicrobiologically. Alternatively or additionally, vesicles, liposomes ormicellar structures are conceivable as primary carriers.

In the case of a particularly advantageous embodiment of the sensorarrangement according to the invention, it is anticipated that theelectrode area exhibits at least one electrically conductive electrode,that an electrically insulating insulation area is provided and that theelectrode concerned is electrically insulated from the measuring medium,from the primary carriers and from the biological units by theinsulation area.

In this case, the electrode is formed by the intermediate substrate areaor by the connecting substrate area, for example, and in particular bythe actual electrode layer. The biomaterial area forms the insulationarea in this case.

The electrode area also advantageously possesses at least one electrode.This can be formed, on the one hand, as such as a mechanically stablematerial area.

On the other hand, the electrode area can also exhibit a carrier whichis formed in particular in the form of a solid body. This function isassumed by the carrier substrate area, for example. It is then possiblefor the electrode to be formed, respectively, as a material area ormaterial layer on a surface area or the surface of this carrier, inparticular in a continuous manner. In this respect, it is anticipated inparticular for the electrode to achieve mechanical stability as a resultof the solid body support provided by the carrier. This procedure hasthe advantage that, if necessary, high-value materials can be appliedonto the carrier as a thin layer, for example, such that the possibilityof a single use sensor electrode device presents itself in an economicoperating respect, which device can be manufactured at affordable pricesand utilised on the market. If necessary, the carrier, in particular theelectrode area, can be recycled in which case, in particular, a replaceinsulation area, e.g. a new thiol layer, may become necessary.

Preferably, the electrode exhibits at least one metallic material or isformed of such a material. In this respect, a chemically inert noblemetal, in particular, preferably gold, is advantageously used. Platinumor silver, in particular, are also conceivable.

Moreover, the use of electrically conductive metal oxides for or in theintermediate substrate area or the connecting substrate area, e.g. ofITO or indium tin oxide, is conceivable. From or with this class ofmaterial, counter-electrodes, which may need to be provided, may bemanufactured.

The carrier for accepting the electrode consequently advantageouslyexhibits an electrically insulating material or is formed of such amaterial. Moreover, or alternatively, it is advantageous for thematerial of the carrier to be essentially chemically inert.Advantageously, glass or the like offers itself as a material. In thisrespect, the shape may that of a panel or such like. The chemicalinertness prevents both a modification of the carrier and acontamination of the measuring medium during the measuring process. As aresult of the selection of an electrically insulating carrier, it isguaranteed that all measuring signals originate essentially from thearea of the electrode.

A possible arrangement of the sensor electrode device is obtained if theelectrode is essentially formed as material layer deposited on thesurface of the carrier. It can also be a vapour deposited or sputteredmaterial layer. The material layer for the formation of the electrodehas a layer thickness of approximately 10 nm to 200 nm, for example.

Between the material layer for the electrode and the surface of thecarrier, an adhesive layer may, if necessary, be of advantage. Onapplying a gold electrode onto glass, in particular, an adhesive layerof chromium or such like present in between is of advantage.Advantageously, the adhesive layer has a relatively low layer thickness,preferably of approximately 5 nm.

To form the capacitive electrode and the insulation of the electrodearea from the measuring medium and/or from the primary carriers, whichis necessary for this purpose, at least one insulating area orbiomaterial area is thus preferably formed as a result of which theelectrode area, in particular the electrode, is essentially electricallyinsulatable in operation, in particular in areas thereof which areprovided for mechanical contact with the measuring medium and/or theprimary carriers during operation.

In a further preferred embodiment of the sensor arrangement or sensorelectrode arrangement according to the invention, the insulation area orbiomaterial area is formed in the form of layers. In this respect, theinsulation area or biomaterial area consists at least partly of asequence of monolayers, the monolayers being formed as spontaneouslyself-organising layers.

In this respect, it is advantageous for a layer of an organic thiocompound to be provided as a sub-layer of the insulation area orbiomaterial area or as a bottom most area, or an area facing towards theelectrode, of the insulation area or the biomaterial area, with a viewto the electrical properties and the electrical insulation, preferablyof a long-chain alkane thiol, in particular of octadecane thiol.

Moreover a layer of an amphiphilic organic compound, in particular of alipid, is provided as top layer of the insulation area or biomaterialarea, as an uppermost area facing away from the electrode or surfacearea of the insulation area.

It can thus be advantageous to form the insulation area or biomaterialarea at least partly in layer form, in particular in multilayer form. Inthis way, the insulation effect is strengthened and the manufacturesimplified. In order to obtain as high a rate of attached and/orarranged primary carriers as possible in the area of the sensorelectrode device, it is anticipated according to a preferred embodimentof the sensor electrode device according to the invention that at leastthe surface area of the insulation area is formed in such a matchedmanner that an attachment and/or arrangement of primary carriers on thesurface area of the insulation area is promoted, in particular in amanner compatible with the surface of the primary carriers. This meansthat, depending on the surface properties of the primary carrier, thesurface area of the insulation area of the sensor electrode device beformed in a correspondingly adjusted manner such that the primarycarriers attach themselves in a favoured manner to the surface area ofthe insulation area and remain there.

With respect to a particularly marked capacitive coupling of the sensorelectrode device during measuring operation, it is therefore anticipatedthat the insulation area is formed at least partly as a single layer,monolayer and/or as a sequence thereof. In this case, the specificarea-related electric capacitance of the electrode boundary layer isparticularly high. The arrangement and formation of the sensorarrangement according to the invention is particularly simple if thelayer or layers of the insulation area are formed as spontaneouslyself-organising layers or as self-assembling layers. In this respect,the tendency and the endeavours of certain essentially liquid startingmaterials or those dissolved in the liquid state to form, on a surface,under the influence of the interaction with the structure of thesurface, spontaneously and in a self-organising manner, an orderedand/or layer-type structure which, under certain circumstances and inthe case of certain classes of substances leads to the formation ofparticularly thin and, if necessary, single-layer layers or monolayers,in particular of molecules, are exploited in an advantageous manner.

In the case of the use of, according to the invention, organic thiocompounds, in particular of alkane thiols, use is made of the fact that,on certain noble metal surfaces, e.g. gold, silver and platinum, it ispossible to form, from an organic solution which contains thecorresponding thio compound in solution, a covalently bonded monolayeron the electrode surface as a result of a specific chemical interactionof the thio group with the surface atoms of the noble metal electrode,which monolayer is capable of forming a hexagonal dense package in thecase of a corresponding geometry of the thio compound, as a result ofwhich a particularly low residual conductivity of the noble metalsurface is achievable with respect to the measuring medium to beprovided.

Correspondingly, it is possible when using metal oxides in the area ofexposed surface areas of the intermediate substrate area or theconnecting substrate area, in particular of indium tin oxide, to makeuse of a correspondingly specific siloxane chemistry for the formationof a covalently bonded sub-layer of the insulation area or thebiomaterial area, in which, as top layer or as uppermost layer and areafacing away from the electrode or surface area of the insulation area orbiomaterial area, a layer of an amphiphilic organic compound, inparticular a lipid and/or the like is provided.

As a result of the procedure in which, as top layer or as uppermost areafacing away from the electrode or surface area of the insulation area orbiomaterial area, a layer of an amphiphilic organic compound, inparticular a lipid and/or the like is provided, a particularlywell-defined arrangement and structurisation of the surface of theinsulation area or biomaterial area is forcefully obtained.

The amphiphilic organic compounds possess at least one area of polarformation such that a certain partial solubility arises in the measuringmedium which, in particular, is of an aqueous nature. On the other hand,amphiphilic organic compounds possess a non-polar or hydrophobic areawhose arrangement in an aqueous measuring medium is less preferred fromthe energy point of view. As a result of this phenomenon, a layerstructure is preferably formed in the case of which the polar orwater-soluble areas of the amphiphilic compounds are allocated to theaqueous measuring medium whereas the non-polar or hydrophobic areas ofthe amphiphilic organic compounds are arranged facing away from theaqueous measuring medium. Consequently, a monolayer can be formed whichforms, in particular, the surface area of the electrode area. This ispreferably done in combination with an alkane thiol monolayer assub-layer such that, at least partly, a double layer of two monolayersis formed as insulation area or biomaterial area.

The sequence of two monolayers thus formed has certain structuralsimilarities to certain membrane structures which are known frombiological systems such that a certain membrane structure can beallocated to the sequence of two monolayers thus formed—namely thealkane thiol monolayer facing towards the electrode and the lipidmonolayer arranged on top. As a result of the basic solid body carrier,this membrane structure is also referred to as solid body supportedmembrane SSM (SSM: solid supported membrane). This SSM membranestructure has particularly advantageous properties with respect to thearrangement and characteristic property of the sensor electrode deviceaccording to the invention, as a capacitively coupled electrode.

The area which is defined by the electrode-insulating and/or coveringlayer of the insulation area or biomaterial area, in particular,exhibits the membrane structure just described in an advantageousmanner. In this respect, it is also advantageous that this membranestructure or SSM has at least in part a specific electric conductivityof approximately G_(m)≈1-100 nS/cm². Moreover, a specific electriccapacitance of approximately C_(m)≈10-1000 nF/cm² is advantageouslypresent. Finally a surface for the membrane structure of approximatelyA≈0.1-50 mm² is provided alternatively or as a supplement.

The high specific capacitance Cm is of particular advantage with respectto an amperometric active principle test to be carried out, in the caseof which initiated electrical actions of the essentially biologicalunits are measured as electric currents, namely as displacement currentsor capacitive currents.

With a view to the signal-to-noise ratio, a corresponding sealantresistance in the area of a few nanosiemens is of particular advantage.

According to another embodiment of the sensor arrangement according tothe invention, this can also be achieved by applying a Teflon layer,e.g. directly onto the metal electrode. Such a procedure is entirelysufficient for potentiometric active principle testing, for example,since, in this case, it is not a high electrical capacitance which isimportant but a high sealing resistance because of the voltagemeasurements.

Particularly simple geometric circumstances arise, in particular with aview to the reproducibility of the measured results, if the carrier, theelectrode and/or the insulation area and/or its surface or boundarysurface areas are formed at least partly in an essentially planarmanner, in particular also at the microscopic level or scale. Theplanarity guarantees that certain field strength effects at the edges ortips which may lead to the breakthrough of the sealing resistance, donot arise. Moreover, with a view to the exchange, in operation, of themeasuring medium to be provided, the advantage of a homogeneous boundarysurface distribution arises. Any possible protuberances or cavitieswould lead to concentration inhomogeneities at the boundary surfacebetween the insulation area and the measuring medium, whichinhomogeneities could possibly have a negative influence on the resultsof detection or measurement achieved. The planarity, in particular ofthe metallic boundary surfaces, can be guaranteed by correspondingmanufacturing processes, e.g. by epitactic growth, annealing or suchlike.

For external contacting of the sensor arrangement, e.g. with an externalmeasuring circuit or the like, a contact area is provided, acorresponding insulation to avoid other short circuits, in particularwith respect to the measuring medium, being formed.

In particular with a view to a high rate of throughput in the case ofactive site and/or active principle tests to be carried outcorrespondingly, it is particularly advantageous if the sensorarrangement according to the invention is formed in such a way that, atleast in operation, it exhibits essentially constant mechanical,electrical and/or structural properties vis-à-vis liquid streams with ahigh flow rate, preferably in the region of approximately v≈0.1-2 m/s,in particular in the region of the membrane structure and/or especiallywith a view to the attachment and/or arrangement of primary carriers.This required and advantageous consistency of the mechanical, electricaland/or structural properties of the sensor arrangement according to theinvention and, in particular, the membrane structure provided therein isobtained inherently as a result of the above-mentioned measures for theformation of the electrode and the insulation layer covering theelectrode, in particular in the form of self-assembling monolayers of analkane thiol on gold with a corresponding monolayer of lipid in anaqueous medium.

Advantageously, the sensor arrangement according to the invention isused with the sensor electrode device described, in a process foramperometric and/or potentiometric, pharmacological active site and/oractive principle testing and in a device for carrying out such aprocess.

In the case of the sensor arrangement according to the invention, aeukaryotic cell, a prokaryotic cell, organelles thereof, a bacterialunit, a viral unit and/or such like and/or components, fragments, inparticular membrane fragments of such like and/or associations thereofin an essentially native and/or modified, in particular purified form orform modified microbiologically and/or by molecular biology are providedas primary carriers respectively.

It is thus conceivable in principle that insulated and whole cells areused as primary carriers of corresponding biological units which can beactivated to an electrical action, irrespective of whether these are ofplant or animal origin. Thus, an examination of entire heart cells, forexample, is possible and conceivable. On the other hand, the examinationof plant cells, for example algae cells or other unicellular organisms,can also be considered. In addition, certain bacteria or viruses can beexamined as a whole. Moreover, it is conceivable to use components orfragments of cells, bacteria or viruses as primary carriers as a resultof specific microbiological or biochemical measures. Also, it isconceivable to use associations of cells, bacteria or such like asprimary carriers and to connect these to the corresponding sensorelectrode device for the formation of a sensor arrangement according tothe invention.

Moreover, the possibility exists according to the invention of using thesuggested primary carriers in their native form or in a modified form.In this respect, eukaryotic cells, prokaryotic cells or bacteria, forexample can be used which have been modified by correspondingpurification, microbiological and/or molecular biology processes inorder to preferably form specific proteins with certain desiredproperties, for example.

Apart from the primary carriers already available in their natural formin the form of cells, bacteria and the like, it is also conceivable toproduce artificial primary carriers in the form of vesicles, liposomes,micellar structures and/or the like, for example. If necessary, theseare then provided and/or enriched with corresponding biological unitswhich can be activated to electrical action. Corresponding processes forthe reconstitution of membrane proteins or such like in vesicles orliposomes are known and can be exploited here in an advantageous mannerin order to create particularly advantageous embodiments of the sensorarrangement according to the invention.

Suitable as essentially biological units are all units which can betriggered into an at least partly electrically produced action. Suchbiological units are conceivable in particular which are activable toperform an at least partial electrogenic and/or electrophoretic chargecarrier transport and/or an at least partial electrogenic orelectrophoretic charge carrier movement and which represent biological,chemical and biochemical units. These are in particular transport unitswhich move charge carriers upon their activation. Components, fragmentsand/or associations of such units, in particular transport units, arealso conceivable.

Membrane proteins, in particular ion pumps, ion channels, transporters,receptors and/or such like offer themselves in particular as biologicalunits. With respect to many of these biological units, findings and/orassumptions exist to the effect that certain processes are associatedwith at least one electrogenic partial step. These electrogenic partialsteps can be associated with an actual substance transport such as inthe case of a channel, an ion pump or certain transporters, for example.However, biological units, in particular membrane proteins, are alsoknown whose electrical activity is not connected with a net materialtransport but rather with a, if necessary reversible, chargedisplacement within the framework of a conformation change or bonding orsuch like. Such electrical activities, too, are measurable, inprinciple, according to the invention as short-term displacementcurrents and/or potential changes.

The biological units, in particular the membrane proteins, can beprovided in essentially their native form and/or in a modified, inparticular purified form or a form modified microbiologically and/or bymolecular biology, respectively. On the one hand, certain nativeproperties, can be tested and pharmacologically investigated in theorganism of existing proteins, for example. On the other hand,modifications initiated by molecular biology or gene technology alsooffer themselves for analysing certain aspects, e.g. the transportationor the pharmacological mode of action of an active principle.

It is particularly advantageous that primary carriers of an essentiallyuniform type of primary carrier are provided in each case. This is ofimportance with regard to as unambiguous as possible as evidence andanalysis of an active substance test and relates to the geometric,physical, chemical, biological and molecular biological properties ofthe primary carrier.

The same also applies to the biological units provided for the primarycarrier, in particular to the membrane proteins or such like. In thiscase, biological units of an essentially uniform type are provided ineach case, in particular with respect to their geometrical, physical,chemical, biological and molecular biological properties. In addition,the biological units should advantageously be approximately uniform withrespect to their orientation and/or with respect to their activatibilityin relation to the primary carrier concerned.

To achieve as high a signal quality as possible, it is advantageous forthe surfaces of the primary carrier and/or the secondary carrier to beformed in such a way that an attachment and/or arrangement of theprimary carriers on the secondary carrier is promoted. In this way, aparticularly high number of attached primary carriers and/or aparticularly close contact of the primary carriers to the secondarycarrier is obtained, on the one hand, as a result of which theelectrical connection and consequently the signal-to-noise ratio areincreased.

The attachment can be controlled e.g. via the so-called lipid-lipidinteraction between the primary carrier, e.g. vesicle, and the secondarycarrier, e.g. lipid thiol SSM. On the other hand, a covalent bond of theprimary carrier to the surface of the secondary carrier is conceivable,e.g. in the form of a biotin-streptavidin scheme or according to themeaning of His-Tag coupling.

In this connection, it is particularly advantageous if the surfaces ofthe primary carriers and of the secondary carrier are formed with anopposite polarity to each other. This promotes the rate of attachment ofthe primary carriers to the secondary carrier and the strength of thecontact between them.

It is particularly advantageous if vesicles or liposomes withessentially the same effect and/or of the same type, preferably of alipid, are provided as primary carriers in and/or on the membrane ofwhich units of essentially one type of membrane protein are embeddedand/or attached in preferably essentially an oriented form.

The sensor arrangement according to the invention is advantageously usedin a process for amperometric and/or potentiometric, in particularpharmacological active site and/or active principle testing and/or in adevice for carrying out such a process.

According to a further aspect of the present invention, a process formanufacturing a biologically compatible or biocompatible sensorelectrode arrangement, in particular a sensor electrode arrangement foramperometric and/or potentiometric, pharmacological active site and/oractive principle testing is created.

In the manufacturing process according to the invention, at least onecarrier substrate area with a top side with a surface area or with a topside surface area is formed. Moreover, at least one intermediatesubstrate area or a connecting substrate area is formed on the surfacearea or the top side surface area of the carrier substrate area or apart thereof, in particular in a structured manner and with a top sidefacing away from the carrier substrate area with a surface area or witha top side surface area. Moreover, a biomaterial area is formed on thetop side surface area of the intermediate substrate area or theconnecting substrate area or a part thereof, in particular in astructured manner, with at least one biologically compatible orbiocompatible material component. According to the invention, thecarrier substrate area with the intermediate substrate area or theconnecting substrate area thereon or the intermediate substrate area orthe connecting substrate area as such and/or a part thereof in each caseare formed in the form or the manner of a wafer element or a printedcircuit. Alternatively or additionally, it is anticipated that thecarrier substrate area with the intermediate substrate area and theconnecting substrate area thereon or the intermediate substrate area orthe connecting substrate area as such and/or a part thereof in each caseare formed as a or with a photolithographically processed structure oras a or with a photographically processed element, as a or with astructure processed by being bonded on or laminated on or as an or withan element processed by being bonded on or laminated on, as a or with astructure processed micromechanically and/or by laser ablation or as anor with an element processed micromechanically and/or by laser ablationand/or a structure processed by printing or as an or with an elementprocessed by printing. This is provided in particular on the carriersubstrate area in each case.

Basic aspects of the manufacturing process according to the inventionconsequently need to be seen in the fact that a carrier substrate area,an intermediate substrate area or a connecting substrate area thereonand a biomaterial area are provided on the surface area of theintermediate substrate area or the connecting substrate area. In thisconnection, it is a further aspect that the carrier substrate area withthe intermediate substrate area or the connecting substrate area thereonor the intermediate substrate area or the connecting substrate area assuch and/or a part thereof in each case are processed in a mannerpossible for wafers or printed circuits in order to achieve aparticularly reliable and cost-effective manufacture, in particular inmass manufacture.

The further manufacturing modes with a view to the provision ofphotolithographically processed structures or elements, bonded-on and/orlaminated-on processed structures or elements, structures or elementsprocessed micromechanically and/or by laser ablation and/or structuresor elements processed by printing need to be additionally oralternatively provided.

The ablation and/or laser ablation takes place, if necessary, with masksupport.

The individual process steps of the manufacturing process according tothe invention and/or their modifications are also carried out in linewith the structural measures described above.

Thus, it is anticipated according to a preferred embodiment of themanufacturing process according to the invention that the carriersubstrate area is formed with a chemically inert, biologically inertand/or essentially electrically insulated material or of such amaterial.

Moreover, it is anticipated alternatively or additionally that thecarrier substrate area is formed with a mechanically flexible materialor of such a material, in particular in the form or in the manner of afilm.

With regard to the structurisation of the biocompatible materialarrangement as sensor electrode arrangement, it is additionally oralternatively provided in the case of an another embodiment of themanufacturing process according to the invention of a sensor electrodearrangement that, for the intermediate substrate area or for theconnecting substrate area, a layer of an electrically conductive metaloxide, for example ITO or indium tin oxide, is formed.

In the case of another embodiment of the manufacturing process accordingto the invention, it is anticipated that, for the intermediate substratearea or for the connecting substrate area, a metallic layer structure isformed on the top side surface or the top side surface area of thecarrier substrate area.

In this connection, it may be anticipated in an advantageous manner thatthe layer structure for the intermediate substrate area or for theconnecting substrate area is formed with at least one or of a primarymetal area arranged bottom most, a subsequent auxiliary layer and anactual electrode layer arranged top most.

In this connection, the primary metal layer or the primary metal area isformed as an alloy barrier and/or diffusion barrier.

It is moreover preferred that the primary metal area is formed with orof copper.

Moreover, it is preferred that the primary metal area is formedphotolithographically. Alternatively or additionally, the possibilityoffers itself to process by bonding on, laminating on, ablation and/orprinting on.

The auxiliary layer is advantageously formed of nickel or containingnickel.

According to a further alternative embodiment of the manufacturingprocess according to the invention, the actual electrode materialarranged top most or the actual electrode layer arranged top most isformed with or of noble metal, preferably with or of gold.

It is particularly preferred that the auxiliary layer and/or the actualelectrode layer arranged top most is formed by electrodeposition.

Alternatively or additionally, micromechanical processing and/or laserablation can be used.

Particularly advantageous properties of the sensor electrode arrangementto be manufactured are obtained if, according to a preferred embodimentof the manufacturing process, the carrier substrate area is formedentirely or partly of a chemically inert, biologically inert materialand/or a material that is at most slightly absorptive vis-à-visproteins, biological and/or chemical active principles.

In the case of a further advantageous embodiment of the manufacturingprocess according to the invention it is anticipated that the carriersubstrate area is formed entirely or partly of PMMA, PTFE, POM, FR4,polyimide such as e.g. PI or Kapton, PEN, PET and/or of a material thatis transparent—particularly in the UV range.

For further flexibilisation and enlargement of the area of use of theproduct to be manufactured according to the meaning of the sensorelectrode arrangement to be manufactured, it is anticipated in the caseof a further advantageous development of the manufacturing processaccording to the invention that a plurality—in particular ofhomogeneous—intermediate substrate areas or connecting substrate areasand/or biomaterial areas is formed. These may be formed in a connectingor in a separate form, in particular with a view to their electricalconnection and/or electrical insulation with and/or from each other.This takes place, in particular, in a laterally separated manner.

In an advantageous manner, the plurality of intermediate substrate areasor connecting substrate areas and/or biomaterial areas is arranged inseries or in matrix form.

According to a further advantageous embodiment of the manufacturingprocess according to the invention, the sensor electrode arrangement isformed as a sensor electrode arrangement for amperometric and/orpotentiometric, pharmacological active site and/or active principletesting.

In this connection, the intermediate substrate area and/or thebiomaterial area are provided in each case as membrane sensor electrodearea or as secondary carrier of the sensor electrode arrangement.

In the case of another embodiment of the process according to theinvention, the intermediate substrate area or the connecting substratearea and the biomaterial area are formed, in each case, as a membranebiosensor electrode area or as a secondary carrier with an electricallyconductive and solid body-type electrode area.

In this connection, a plurality of primary carriers is provided in theimmediate spatial vicinity of the secondary carriers or the secondarycarrier. In this case, the primary carriers contain, biological unitsactivable into electrical action, in particular membrane proteins.

According to the invention, a eukaryotic cell, a prokaryotic cell, abacterium, a virus or components, in particular membrane fragments orassociations thereof in the native form or in the modified form, inparticular in the purified, microbiological form and/or form modified bymolecular biology are provided as primary carrier in each case.Alternatively or additionally, a vesicle, a liposome or a cellularstructure are provided as primary carrier.

Moreover, it may be anticipated that the intermediate substrate area orthe connecting substrate area is provided as at least one electricallyconductive electrode of the electrode area, that the biomaterial area isprovided as an electrically insulated insulation area and that, inoperation, the electrode concerned is electrically insulated by thebiomaterial area or the insulation area from a measuring medium, fromthe primary carriers and from the biological units.

According to a further embodiment of the process according to theinvention, it is anticipated that the biomaterial area or insulationarea is formed in layers, that the insulation area is formed at leastpartly of a sequence of monolayers and/or that the monolayers are formedas spontaneously self-organising layers.

In the case of another embodiment of the process according to theinvention, it is anticipated that, as a sub-layer of the biomaterialarea or the insulation area, a layer of an organic thio compound isprovided as a bottom most area of the insulation area facing towards theelectrode, preferably of a long-chain alkane thiol, in particular ofoctadecane thiol, and that, as top layer of the biomaterial area or theinsulation area, a layer of an amphiphilic organic compound, inparticular of a lipid, is provided as uppermost area facing away fromthe electrode or surface area of the insulation area.

In the case of a further advantageous embodiment of the manufacturingprocess according to the invention, it is anticipated that the area ofthe biomaterial area or the insulation area insulating and covering theelectrode is formed with a membrane structure with a surface ofapproximately A≈0.1-50 mm² and with a specific electric conductivity ofapproximately G_(m)≈1-100 nS/cm² and/or with a specific capacitance ofapproximately C_(m)≈10-1000 nF/cm².

According to a further preferred embodiment of the manufacturing processaccording to the invention, it is anticipated that a biological unit isprovided which is formed to be activable to an electrogenic chargecarrier movement, in particular to an electrogenic charge carriertransport.

Additionally or alternatively, it is anticipated that a membraneprotein, in particular an ion pump, an ion channel, a transporter or areceptor or a component or an association thereof is provided asbiological unit in each case.

Moreover, it is preferred alternatively or additionally that thebiological unit is provided in native form or in a modified form, inparticular in a purified, microbiologically modified form and/or a formmodified by molecular biology.

In a further advantageous development of the manufacturing processaccording to the invention, it is anticipated that the surface of theprimary carriers and the surface of the secondary carriers are formedwith opposite polarity or oppositely charged to each other and/or that,between the surface of the primary carriers and the surface of thesecondary carrier, a connection in the manner of a chemical bond isformed, in particular via a His-Tag coupling or a streptavidin-biotincoupling or the like.

These and other aspects of the present invention result, in other words,also from the following remarks:

The invention relates not only to corresponding structures but also to aprocess for the manufacture of electrically insulating, extremely thinlayers as biocompatible areas or material areas, in particular onprinted circuit boards or the like and to their use as sensor elements,in particular for single use.

In the field of bioanalysis, it is desirable in certain cases to havebiocompatible surfaces available which are suitable for the absorptiveattachment of biological membranes, membrane fragments or of artificiallipid double layers. The task, on which the invention described hereinis based, consisted of manufacturing such surfaces in as cost-effectivea manner as possible without suffering restrictions in functionality.

Methods are known which operate on the basis of optical measured valueson biocompatible layers, so-called biacore measurements, for example,according to the principle of surface plasmon resonance or measurementsof the load increase change using the quartz microbalance. In othercases, electrical properties of the attached, often protein-containingvesicles, cells or membrane fragments are to be detected.

Frequently, substrates or carrier substrate areas of mica, glass orquartz can be used which are coated e.g. with gold, by thin layertechnique.

Occasionally, the selection of the substrate needs to be made on thebasis of the specific properties of the substrate, e.g. glass, becauseof its transmittance in the area of visible light and because of itsrefractive index, quartz as a result of its ability to be induced tooscillation in the condenser field. Occasionally, glass is used becauseof its chemical inertness and the possibility of lithographicstructuring of the gold layer down into the microstructure region.

Providing mica, glass and/or quartz with the intermediate substrate areaor connecting substrate area by epitaxial growth, by vapour depositionand/or sputtering, for example, is meaningful and anticipated accordingto the invention in those cases where the controllability, highdefinition, high value and/or planarity of the intermediate substratearea or connecting substrate area and/or the corresponding surface areasare of importance.

It is also conceivable that areas can be produced on gold surfaces whichare capable of providing specific bonding for target molecules.

The manufacture of structured biocompatible areas on gold surfaceswhich, in turn, have been applied onto glass substrates is complicatedand costly under certain circumstances. Moreover, glass is fragile andmay form, under certain circumstances, sharp edges capable of causinginjuries.

These properties lead to the replacement according to the invention ofglass substrates, mica or quartz as carrier for biocompatible areasaccording to the meaning of the invention by other materials and/orcoating techniques other than epitaxial growing, vapour depositionand/or sputtering.

The use of a self-organising monolayer, a self-assembled monolayer or anSAM does not lead to the desired or necessary electrical properties andonly partly to the ability of the surface to adsorb vesicles, cells ormembrane fragments.

It is a core idea of this invention to produce, on printed circuitboards or such like producible cost-effectively in very large numbers,for example, biocompatible areas which provide a very low electricconductivity between the actual electrode or intermediate substrate andthe surroundings and exhibit suitable adsorption properties with respectto cells, cell membrane fragments, liposomes or such like.

The structurisation of the printed circuit boards or such like takesplace by selective or structured coating of a primary metal layer, e.g.a copper layer, for example, by subsequently selectively removing theprimary metal layer, e.g. by wet-chemical etching, and by subsequentfinishing, e.g. by gold plating.

These techniques permit the manufacture of very large numbers of items,the costs being lower by a multiple than those in the case of glasssubstrates. Moreover, biocompatible areas serving as biosensor can bemanufactured in the immediate spatial vicinity to amplifier devices onwafers such that noise and interference can be considerably reduced. Thesubstrates used for the manufacture are highly stable and, optionally,do not have sharp edges. They are therefore highly suitable formanufacturing disposables.

A copper-coated printed circuit board—e.g. with 17 μm copper—, forexample, is coated with photoresist. A layout is transferred onto thephotoresist by light exposure. The photoresist is developed and removedspecifically in the areas not exposed to light, the copper layer beingexposed in those areas. The copper layer is removed at the exposedsites. The remaining resist residues are also removed. Nickel, forexample, is electrodeposited onto the free copper structures. Gold, forexample, is electrodeposited onto the nickel layer thus obtained.

An alkane thiol monolayer is produced on the gold layer asself-assembled monolayer or SAM. By adding lipid-containing solution, ahybrid lipid layer is produced on the SAM in a manner analogous to alipid double layer. This hybrid lipid layer permits the stableadsorption e.g. of membrane fragments of biological membranes, cellfragments, vesicles and liposomes.

By integrating the printed circuit boards or the like into an electricamplifier circuit and by integrating the biocompatible area into a flowcell and by introducing an Ag/AgCl reference electrode into thefluid-coupled system and by attaching membrane fragments withelectrogenic membrane proteins, the modified printed circuit boards canbe used as biosensors.

Further aspects of the present invention arise as follows:

Frequently, glass and/or a complicated technical process are used inorder to obtain thin, very high quality intermediate substrate areas,connecting substrate areas and/or actual electrode layers according tothe meaning of the invention, in particular gold layers, with it. Thebasic idea has been that a slightly rough surface is particularlyadvantageous.

This process may be uneconomical. As an alternative, it is thus possibleto use intermediate substrate areas, connecting substrate areas and/oractual electrode layers, i.e. gold layers, for example, andcorresponding surface areas of comparatively extremely poor quality withrespect to visible granularity under the light microscope, layerthickness of several micrometers, beads, scratches etc, for example.

However, it has been found that essential characteristics of themembrane biosensor electrode area, i.e. the SSM, are retained such thatthe materials mica, glass, quartz and/or the application orstructurisation by epitaxial growing, by vapour deposition and/or bysputtering are not necessarily required.

Consequently, fields of application for these comparatively low valuebut also cheap electrodes thus specifically arise in an advantageousmanner.

Consequently, comprehensive extensions of the comparatively complexarrangements and the corresponding manufacturing processes thus ariseaccording to the invention.

For this reason, too, the possibility offers itself to structurisegold-vapour deposition treated films, for example, consisting ofpolyimid or PEN, for example, by laser ablation, the laser beam beingpassed through a mask. The film may be drawn from a roll and structuredin a continuous process.

BRIEF DESCRIPTION OF THE DRAWINGS

In the following, the present invention is explained in further detailby way of a diagrammatic representation based on preferred practicalexamples.

FIG. 1 shows a diagrammatic and sectional side view of an embodiment ofthe biocompatible sensor electrode arrangement according to theinvention.

FIGS. 2A, B show a diagrammatic top view and/or a sectional side view ofanother embodiment of the biocompatible sensor electrode arrangementaccording to the invention.

FIG. 3 shows a diagrammatic top view of a further embodiment of thebiocompatible sensor electrode arrangement according to the inventionwith a plurality of individual electrodes.

FIG. 4 shows a diagrammatic and partial sectional side view of anotherembodiment of the biocompatible sensor electrode arrangement accordingto the invention with a vesicle as primary carrier and its use in ameasuring device.

FIG. 5 shows a further embodiment of the biocompatible sensor electrodearrangement according to the invention with a membrane fragment as aprimary carrier.

DETAILED DESCRIPTION OF THE INVENTION

In the following the same references indicate the same, identical oridentically acting structures or elements. A detailed description willtherefore not be repeated each time they occur.

FIG. 1 shows a diagrammatic and sectional side view of a firstembodiment of the biocompatible sensor electrode arrangement 1 accordingto the invention.

This first embodiment of the biocompatible sensor electrode arrangement1 according to the invention exhibits a carrier substrate area 22 or acarrier substrate 22 with a top side surface area 22 a on which theconnecting intermediate substrate area 26 or the connecting substratearea 26 is provided in the form of a layered metal structure, namelywith a primary metal area 26-1, of copper in this case, an auxiliarylayer 26-2, e.g. of nickel in this case, which serves as diffusionbarrier and alloy formation barrier, as well as an actual electrodelayer 26-3, of gold in this case.

By a specific chemical interaction with the actual electrode layer 26-3,a biomaterial layer 24 or a biomaterial area 24 is immobilised on thetop side surface area 26 a of the connecting substrate area 26. Thisbiomaterial area 24 serves as insulation area 24 for the sensorelectrode arrangement 1 according to the invention and consists of alayered sequence of self-organising monolayers 24 a and 24 b, namely ofa sub-layer arranged bottom most in the form of an alkane thiolmonolayer 24 b which is connected via the specific thiol goldinteraction or SH—Au interaction, and a lipid monolayer 24 a provideduppermost. By means of this arrangement, a membrane biosensor electrodedevice M or 20 with a solid body-supported membrane SSM is formed.

FIGS. 2A and 2B show a diagrammatic top view and/or a diagrammatic andsectional side view of another embodiment of the biocompatible sensorelectrode arrangement 1 according to the invention. In this case, aprocessed counter-electrode device 46 is also shown in the top view ofFIG. 2A, which device, however, was left out from the side view of FIG.2B. This counter-electrode 46 can also consist of ITO or indium tinoxide and assume alternative embodiments.

FIG. 3 shows, by way of a diagrammatic top view, an embodiment of thesensor electrode arrangement 1 according to the invention on which sixindividual electrodes 26 with corresponding supply leads 29 are formedon the upper surface 22 a of the carrier substrate 22. The individualelectrodes 26 with their corresponding terminal leads 29 are formed inan essentially identical manner, at least insofar as the manufacturingtolerances allow.

All characteristic properties relating to the mesoscopic or microscopestructure of the surface of the membrane biosensor electrode area M, thesecondary carrier 20 and, in particular, the respective allocatedelectrodes 26 can also be seen in the representation of the followingFIGS. 4 and 5. All the characteristic properties illustrated therein areapplicable in any random combination to the structures described abovein FIG. 1 to 3.

FIG. 4 shows a diagrammatic and partly sectional side view of a furtherembodiment of the sensor arrangement 1 according to the invention and acorresponding device for amperometric and/or potentiometricpharmacological active principle testing.

A measuring chamber 50 in the form of an essentially closed vesselforms, together with an exchanger/mixing device 60 in the form of aperfuser system or a pump facility, for example, a closed liquidcircuit. Communication of the liquid serving as measuring medium 30 iseffected via corresponding feed and discharge devices 51 and/or 52. Themeasuring medium 30 can be an aqueous electrolyte solution in this casewhich exhibits certain ion moieties, a given temperature, a specific pHetc. Moreover, specific substrate substances S and/or specific activeprinciples W are, if necessary, contained in the measuring medium 30 orthey are added in later process steps through the exchange/mixing device60.

In the measuring area 50, a sensor arrangement 1 according to theinvention is provided. The sensor arrangement 1 consists of primarycarriers 10 which are attached to the surface area 24 a of the sensorelectrode device 20 serving as secondary carrier.

In the practical example shown in FIG. 4 in diagrammatic form not trueto scale, only a single primary carrier 10 is shown. This consists of alipid vesicle or liposome in the form of a lipid double layer or lipidmembrane 11 formed as an essentially hollow closed sphere. In this lipiddouble layer 11 of the vesicle serving as primary carrier 10, a membraneprotein is embedded in a manner penetrating through the membrane asessentially biological unit 12.

By converting a substrate S present in the measuring medium 30 into aconverted substrate S′, certain processes are initiated in the membraneprotein 12 which, in the case shown in FIG. 1, leads to a substancetransport of a species Q from the extra-vesicular side or outside 10 aof the vesicle 10 to the intravesicular side or inside 10 b of thevesicle 10. If the species Q has an electric charge, the transportationof the species Q from side 10 a to side 10 b leads to a net chargetransportation which corresponds to an electric current from the outside10 a of the vesicle 10 to the inside 10 b of the vesicle 10.

Into each vesicle 10, a multiplicity of essentially identical membraneprotein molecules 12 are incorporated in essentially the sameorientation into membrane 11 of the vesicle 10 as a rule and on the onehand. If these are essentially simultaneously activated—e.g. by aconcentration jump, initiated by mixing, in the concentration of thesubstrate S of a non-activating measuring medium N, 30 without substrateS to an activating measuring medium A, 30 with substrate S—this leads toa measurable electric current.

This charge carrier transportation is measurable because a multiplicityof primary carriers 10 or vesicles are attached to the surface 24 a ofthe sensor electrode device 20 such that, on activation of amultiplicity of protein molecules 12 in a multiplicity of vesicles infront of the surface 24 a of the sensor electrode device 20, a spatialcharge of a certain polarity is formed. This spatial charge then actsonto the electrode 26 which, in the case shown in FIG. 1, is vapourdeposited onto a carrier 22 of glass in the form of a gold layer andcovered by a double layer, serving as insulation area 24, of a bottomlayer 24 b and a top layer 24 a serving as surface and electricallyinsulated vis-à-vis the measuring medium 30.

The surface or upper layer 24 a of the insulation area 24 is a lipidmonolayer, for example, which is compatible with the lipid double layer11 of the vesicle 10 which monolayer is formed by means of aself-assembly process on an alkane thiol monolayer forming the bottomlayer 24 b in such a way that the sequence of the layers 24 b and 24 a,namely the sequence of an alkane thiol monolayer and a lipid monolayer,forms a membrane structure SSM as electrode 26 on a gold substrateformed in the manner of a solid body, which membrane structure is alsoreferred to as solid supported membrane (SSM).

The sensor arrangement 1 and, in particular, the sensor electrode device20 is connected to a data acquisition/control device 40 via a connectingline 48 i. This device is equipped with a measuring device 44 in whichan electric current I(t) or an electric voltage U(t) can be measured asa function of time. Moreover, an amplifier device 42 is anticipated inwhich the measuring signals are filtered and/or amplified. Via a controlline 48 s, the active principle testing is controlled by controlling theexchange/measuring device 60. Via a further line 48 o, the electriccircuit is closed by a counter-electrode 46, e.g. in the form of a Pt/Ptelectrode or by an Ag/AgCl electrode. Insulations 28, 27 and 47 preventshort circuits of the SSM and/or the counter-electrode 46 vis-à-vis themeasuring medium 30.

FIG. 5 shows a diagrammatic and partly sectional side view of anembodiment of the sensor arrangement 1 according to the invention in thecase of which a membrane fragment 10 is provided as primary carrier 10instead of a vesicle or liposome, into which fragment a membrane proteinis embedded as biological unit 12 in an oriented manner. With respect tothe embodiment of FIG. 5, it should be noted that the representation isnot true to scale and on the other hand, a large plurality of membranefragments are, as a rule, attached or adsorbed simultaneously to the SSMor the surface 24 a of the sensor electrode device 20 serving assecondary carrier.

Here, too, it is shown that, by converting the substrate S provided inthe measuring medium 30 into a converted substrate S′, a substancetransport of the species Q from one side 10 a of the membrane fragment10 to the opposite side 10 b takes place which can be detected via thecorresponding net charge transport and the displacement currentconnected therewith as a function of the time.

The invention has been described with particular reference to thepreferred embodiments thereof, but it should be understood thatvariations and modifications within the spirit and scope of theinvention may occur to those skilled in the art to which the inventionpertains.

1. A biocompatible sensor electrode arrangement comprising: at least onecarrier substrate area having a top side with a surface area; at leastone intermediate substrate area formed on the surface area of thecarrier substrate area or a part thereof in a structured manner andformed with a top side facing away from the carrier substrate area witha surface area; and a biomaterial area formed on the surface area of theintermediate substrate area or a part thereof in a structured manner,with at least one biologically compatible material component, thecarrier substrate with the intermediate substrate area thereon or theintermediate substrate area as such or a part thereof in each case beingformed in the form or the manner of a wafer element or a printedcircuit; the intermediate substrate area being provided as at least oneelectrically conductive electrode of the electrode area; the biomaterialarea being provided as an electrically insulating insulation area; andin operation, the electrode concerned being electrically insulated bythe biomaterial area from a measuring medium, from the primary carriersand from the biological units; the biomaterial area being formed aslayers; the biomaterial area being formed at least partly of a sequenceof monolayers, comprising a sub-layer and a top layer; the monolayersbeing formed as spontaneously self-organising layers; as said sub-layerof the biomaterial area, a layer of a long-chain alkane thiol beingprovided as bottom most area facing towards the electrode of thebiomaterial area; and as said top layer of the biomaterial area, a layerof a lipid being provided as an uppermost area facing away from theelectrode or surface area of the biomaterial area.
 2. The sensorelectrode arrangement according to claim 1, wherein the carriersubstrate area exhibits a chemically inert, biologically inert andelectrically insulating material or is formed of such a material.
 3. Thesensor electrode arrangement according claim 1, wherein the carriersubstrate area comprises a mechanically flexible material or is formedas such.
 4. The sensor electrode arrangement according to claim 1,wherein a metallic layer structure is formed on the surface of thecarrier substrate area for the intermediate substrate area.
 5. Thesensor electrode arrangement according to claim 4, wherein the layerstructure for the intermediate substrate area comprises at least one orof at least one primary metal area arranged bottom most, a subsequentauxiliary layer and an actual electrode layer arranged top most.
 6. Thesensor electrode arrangement according to claim 5, wherein the primarymetal area is formed with or of copper.
 7. The sensor electrodearrangement according to claim 5, wherein the primary metal area isformed by a process selected from the group consisting ofphotolithographically, bonded on, laminated on and printed on.
 8. Thesensor electrode arrangement according to claim 5, wherein the auxiliarylayer is formed with or of nickel.
 9. The sensor electrode arrangementaccording to claim 5, wherein the actual electrode layer arrangeduppermost is formed with or of a noble metal.
 10. The sensor electrodearrangement according to claim 5, wherein at least one of the auxiliarylayer and the actual electrode layer arranged uppermost are formed byelectrodeposition.
 11. The sensor electrode arrangement according toclaim 1, wherein the carrier substrate area is formed entirely or partlyof a chemically inert, biologically inert material and a material atmost slightly absorptive vis-à-vis proteins, biologically and chemicallyactive principles.
 12. The sensor electrode arrangement according toclaim 1, wherein the carrier substrate area is formed with or of amaterial selected from the group consisting of PMMA, PTFE, POM, FR4,polyimide, PI, Kapton, PEN, PET and materials transparent in the UVrange.
 13. The sensor electrode arrangement according to claim 1,wherein a plurality of identical intermediate substrate areas andbiomaterial areas is formed electrically insulated from each other andare laterally arranged side by side on the carrier substrate area. 14.The sensor electrode arrangement according to claim 13, wherein theplurality of intermediate substrate areas and biomaterial areas arearranged in sequence or in matrix form.
 15. The sensor electrodearrangement according to claim 1, wherein said sensor electrodearrangement is formed as a sensor electrode arrangement for at least oneof amperometric, potentiometric, pharmacological active site and activeprinciple testing.
 16. The sensor electrode arrangement according toclaim 1, wherein the intermediate substrate area and the biomaterialarea are each provided in a form selected from the group consisting of amembrane biosensor electrode area and a secondary carrier of the sensorelectrode arrangement.
 17. The sensor electrode arrangement according toclaim 1, wherein the intermediate substrate area and the biomaterialarea are each provided in a form selected from the group consisting of amembrane biosensor electrode area and a secondary carrier with anelectrically conductive and solid body-type electrode area.
 18. Thesensor electrode arrangement according to claim 17, wherein a pluralityof primary carriers is provided in immediate spatial vicinity of thesecondary carrier, the primary carriers being activable to electronicaction and biological action.
 19. The sensor electrode arrangementaccording to claim 18, wherein as primary carrier, a primary carrierfrom the group is provided comprising a eukaryotic cell, a prokaryoticcell, a bacterium, a virus, components, membrane fragments, orassociations thereof in the native form or in a modified form or asprimary carrier, a primary carrier of the group is provided comprising avesicle, a liposome or a micellar structure.
 20. The sensor electrodearrangement according to claim 15, wherein the area insulating andcovering the electrode, of the biomaterial area or the insulation areacomprises a membrane structure with a surface of approximately A≈0.1-50mm² and with a specific electric conductivity of approximatelyG_(m)≈1-100 nS/cm² and/or with a specific capacitance of approximatelyC_(m)≈10-1000 nF/cm².
 21. The sensor electrode arrangement according toclaim 15, further comprising a biological unit activable to electrogeniccharge carrier movement or to electrogenic charge carriertransportation.
 22. The sensor electrode arrangement according to claim15, further comprising a biological unit selected from the groupconsisting of a membrane protein, an ion pump, an ion channel, atransporter, a receptor, a component and an association thereof.
 23. Thesensor electrode arrangement according to claim 22, wherein thebiological unit is provided in the native form or in a form selectedfrom the group consisting of a modified form, a purified form, a formmodified microbiologically and a form modified by molecular biology. 24.The sensor electrode arrangement according to claim 15, wherein, thesurface of the primary carriers and the surface of the secondary carriercomprise an opposite polarity or charge or a connection of the type of achemical bond being formed via a His-Tag coupling or a streptavidinbiotin coupling, between the surface of the primary carriers and thesurface of the secondary carrier.
 25. A process for manfacturing abiocompatible sensor electrode arrangement comprising the steps of:forming at least one carrier substrate area with a top side having asurface area; forming at least one intermediate substrate area on thesurface area of the carrier substrate area or a part thereof in astructured manner and with a top side facing away from the carriersubstrate area with a surface area; and forming a biomaterial area onthe surface area of the intermediate substrate area or a part thereof ina structured manner with at least one biologically compatible materialcomponent; the carrier substrate area with the intermediate substratearea thereon or the intermediate substrate area as such or a partthereof being formed in the form or in the manner of selected from thegroup consisting of a wafer element and a printed circuit; theintermediate substrate area being provided as at least one electricallyconductive electrode of the electrode area; the biomaterial area beingprovided as an electrically insulating insulation area; and in operationthe biomaterial area electrically insulating the electrode from ameasuring medium, from the primary carriers and from the biologicalunits; the biomaterial area being formed as layers; the biomaterial areabeing formed at least partly of a sequence of monolayers, comprising asub-layer and a top layer; the monolayers being formed as spontaneouslyself-organising layers; as said sub-layer of the biomaterial area,providing a layer of a long-chain alkane thiol as a bottom most areafacing towards the electrode of the biomaterial area; and as said toplayer of the biomaterial area, providing a layer of a lipid as anuppermost area facing away from the electrode or surface area of thebiomaterial area.
 26. The process according to claim 25, comprising thesteps of forming the carrier substrate with the intermediate substratearea thereon or the intermediate substrate area as such or a partthereof: as a or with a photolithographically processed structure or asa or with a photographically processed element; as a or with a structurebeing bonded on or laminated on or as an or with an element processed bybeing bonded on or laminated on; as a or with a structure processed byat least one of micromechanically and laser ablation or as an or with anelement processed by at least one of micromechanically and laserablation; and/or as a or with a structure processed by printing or as anor with an element processed by printing on the carrier substrate. 27.The process according to claim 25, wherein the carrier substrate area isformed with or of a material selected from the group consisting of achemically inert material, a biologically inert material and anelectrically insulating material.
 28. The process according to claim 25,wherein the carrier substrate area is formed with a mechanicallyflexible material or of such a material.
 29. The process according toclaim 25, further comprising the step of forming a metallic layerstructure on the top side surface of the carrier substrate area for theintermediate substrate area.
 30. The process according to claim 29,wherein the layer structure for the intermediate substrate area or forthe connecting substrate layer is formed with at least one or of atleast one primary metal area arranged bottom most, a subsequentauxiliary layer as an alloy and/or diffusion barrier and an actualelectrode layer arranged top most.
 31. The process according to claim30, wherein the primary metal area comprises copper.
 32. The processaccording to claim 31, comprising the step of forming primary metal areaby a process selected from the group consisting ofphotolithographically, bonded on, laminated on and printed on.
 33. Theprocess according to claim 32, wherein the auxiliary layer comprisesnickel.
 34. The process according to claim 30, wherein the actualelectrode layer arranged top most comprises a noble metal.
 35. Theprocess according to claim 30, wherein at least one of the auxiliarylayer and the actual electrode layer arranged top most are formed byelectrodeposition.
 36. The process according to claim 25, wherein to thecarrier substrate area is formed entirely or partly of at least onematerial selected from the group consisting of a chemically inertmaterial, a biologically inert material and a material at most slightlyabsorptive vis-à-vis proteins, biologically and/or chemically activeprinciples.
 37. The process according to claim 25, wherein the carriersubstrate area is formed entirely or partially of a material selectedfrom the group consisting of PMMA, PTFE, POM, FR4, polyimide, PEN, PETand a material which is transparent in the UV range.
 38. The processaccording to claim 25, comprising the step of forming a plurality ofintermediate substrate areas or connecting substrate areas and/orbiomaterial areas in a connected or in a separated form.
 39. The processaccording to claim 38, wherein the plurality of the intermediatesubstrate areas and/or biomaterial areas are arranged in sequence or inmatrix form.
 40. The process according to claim 25, comprising the stepof forming the sensor electrode arrangement as a sensor electrodearrangement for amperometric and/or potentiometric, pharmacologicalactive site and/or active principle testing.
 41. The process accordingto claim 40, further comprising the step of providing at least one ofthe intermediate substrate area and the biomaterial area in a formselected from the group consisting of a membrane biosensor electrodearea and a secondary carrier of the sensor electrode arrangement. 42.The process according to claim 41, further comprising the step offorming the intermediate substrate area and the biomaterial area in aform selected from the group consisting of a membrane biosensorelectrode area and a secondary carrier with an electrically conductiveand solid body-type electrode area.
 43. The process according to claim42, further comprising the step of providing a plurality of primarycarriers in immediate spatial vicinity of the secondary carrier, theprimary carriers containing units which are activable to electronicaction and biological action.
 44. The process according to claim 43,further comprising the step of providing said primary carriers as aprimary carrier comprising a eukaryotic cell, a prokaryotic cell, abacterium, a virus or components, membrane fragments, or associationsthereof in the native form or in a modified, purified form or a formmodified microbiologically or by molecular biology or in which, asprimary carrier, a primary carrier comprising a vesicle, a liposome or amicellar structure.
 45. The process according to claim 39, comprisingthe step of forming the area, insulating and covering the electrode, ofthe biomaterial area or the insulation area with a membrane structure(SSM) with a surface of approximately A≈0.1-50 mm² and with a specificelectric conductivity of approximately G_(m)≈1-100 nS/cm² and/or with aspecific capacitance of approximately C_(m)≈10-1000 nF/cm².
 46. Theprocess according to claim 39, comprising the step of providingbiological unit which is activable to electrogenic charge carriermovement.
 47. The process according to claim 39, comprising the step ofproviding a membrane protein as a biological unit.
 48. The processaccording to claim 47, comprising the step of providing the biologicalunit in a form selected from the group consisting of the native form anda modified form.
 49. The process according to claim 39, comprising atleast one of the following steps of forming the surface of the primarycarriers and the surface of the secondary carrier formed with anopposite polarity or charge and forming a chemical bond connectionbetween the surface of the primary carriers and the surface of thesecondary carrier.
 50. The sensor electrode arrangement according toclaim 3, wherein the carrier substrate area is in the form or the mannerof a film.
 51. The sensor electrode arrangement according to claim 5,wherein said subsequent auxiliary layer is at least one of an alloy anda diffusion barrier.
 52. The sensor electrode arrangement according toclaim 9, wherein said noble metal is gold.
 53. The sensor electrodearrangement according to claim 18, wherein said primary carriers areactivatable to membrane proteins.
 54. The sensor electrode arrangementaccording to claim 19, wherein as primary carrier, said primary carrierfrom the group is provided in a form selected from the group consistingof the purified form and a form modified microbiologically and bymolecular biology
 55. The process according to claim 28, wherein thecarrier substrate area is in the form or manner of a film.
 56. Theprocess according to claim 34, wherein said noble metal is gold.
 57. Theprocess according to claim 37, wherein said polyimide is selected fromthe group consisting of PI and Kapton.
 58. The process according toclaim 38, comprising the step of forming said plurality of intermediatesubstrate areas or connecting substrate areas and/or biomaterial areasin a form electrically insulated from each other, wherein saidintermediate substrate areas are identical.
 59. The process according toclaim 43, wherein said primary carriers containing units are activableto biological membrane proteins.
 60. The process according to claim 46,comprising the step of providing said biological unit which is activableto electrogenic charge carrier transportation.
 61. The process accordingto claim 47, wherein said membrane protein is selected from the groupconsisting of an ion pump, an ion channel, a transporter, a receptor anda component or an association thereof.
 62. The process according toclaim 48, comprising the step of providing the biological unit in a formselected from the group consisting of a purified form, a form modifiedmicrobiologically and a form modified by molecular biology.
 63. Theprocess according to claim 49, wherein said chemical bond is selectedfrom the group consisting of a His-Tag coupling and a streptavidinbiotin coupling.